The fields of forensics, paternity testing, tissue typing, and personalized medicine routinely use DNA-based techniques for identity determinations, genotyping, phenotypic prediction, and in the prediction and/or prevention of disease. DNA typing involves the analysis of alleles of genomic DNA with characteristics of interest, commonly referred to as “markers.” Most typing methods in use today are specifically designed to detect and analyze differences in the length and/or sequence of one or more regions of DNA markers known to appear in at least two different forms in a population. Such length and/or sequence variation is referred to as “polymorphism.” Any region (i.e., “locus”) of DNA in which such a variation occurs is referred to as a “polymorphic locus.”
In recent years, the discovery and development of polymorphic short tandem repeats (STRs) as genetic markers has played an important role in DNA typing. STRs have become the primary means for human identity and forensic DNA testing. In 2005, the European forensic community first published recommendations on the development of new multiplexes to provide greater discrimination power and enhanced performance on the increasing number of difficult samples encountered by forensic laboratories. Recently, the number of loci was expanded, referred to as the European Standard Set of Loci (ESSL), to include the SE33 locus for those European countries that chose to analyze this highly informative marker. The German National DNA database has included SE33 in the standard set of loci and SE33 is also profiled by neighbouring countries for data sharing purposes. SE33 is also known as ACTBP2 for β-actin pseudogene or as HUMACTBP2 and is located at 6q14. It has a high level of short tandem repeat (STR) complexity contributing to multiple length and sequence micorvariants that often create difficulties in identifying stable primer-binding sites and in primer design. With its high mutation rate and complicated short tandem repeat sequence SE33 is also highly discriminatory and useful in forensics, disaster victim identification, missing persons investigations and complex kinship analysis.
Accurate DNA analysis has both identified missing persons and exonerated the innocent. The adoption of DNA test results has established DNA-based methodologies as a standard investigative, diagnostic or prognostic tool depending on the application. Matching DNA profiles produced from existing commercial STR assays with improved STR assays provides continuity and comparability of the DNA profiles within and between databases. An alteration in the DNA sequence due to, for example, an unknown mutation, polymorphism or re-arrangement, can result in allelic dropout (the failure or significantly reduced amplification of a target nucleic acid) or mobility shift (discordance of results for the same sample in relation to an allelic ladder when analyzing DNA fragment length on an electrophoresis instrument). The identification of such alterations can be useful in the continued effort to maintain the sensitivity, specificity, quality and reliability of DNA-based technologies. The occurrence of a shift in mobility or allelic dropout in new STR assays can make DNA profile matching within and between databases difficult or imprecise. Thus, careful design of new assays such that all potential amplification products are detected in as large a portion of the population as possible remains an ongoing concern when developing new STR assays. Therefore, there exists a need in the art, to improve DNA-based technologies based on the discovery of new variations in human DNA sequences.